B-cell non-Hodgkin lymphomas (NHL) are common lymphoid malignancies in which the infiltration of T lymphocytes correlates with the outcome of patients. Despite extensive studies on anti-tumor immunity, the pathophysiological significance of infiltrating T cells in B-cell NHL remains poorly understood. Recent studies have suggested that CD4+CD25+ regulatory T (Treg) cells are involved in the regulation of anti-tumor immunity by inducing peripheral tolerance to tumor specific antigens. However, there are little data regarding the effect of Treg cells on tumor-specific T cell immunity in B-cell NHL and subsequently on the malignant B-cell growth. In preliminary studies supported by a SPORE development award, we have identified a subset of CD4+CD25+ T cells with a Treg cell phenotype that are present in B-cell NHL.In addition, we find that these Treg cells have the ability to suppress tumor-infiltrating T cells in B-cell NHL and that they migrate in response to factors such as CCL22 produced by the malignant B-cells. Our central hypothesis is that tumor Treg cells contribute to the growth of malignant lymphoma B cells by suppressing tumor-infiltrating T cells and that malignant B-cells play an active role by selectively recruiting Treg cells to the areas of B-cell NHL. We therefore propose to firstly determine the mechanism by which these Treg cells are recruited to the malignant B-cell microenvironment in non-Hodgkin lymphoma and to discover whether they gain suppressive activity when present in the tumor microenvironment (Aim 1). Secondly, we will assess whether malignant B-cells interact directly with Treg cells in the tumor microenvironment and thereby orchestrate tolerance to their presence (Aim 2). Thirdly, we will establish whether depletion of intratumoral Treg cells, and inhibition of malignant B-cells to decrease Treg cell recruitment, will result in clinical benefit for patients with B-cell NHL (Aim 3). We anticipate that the proposed research will provide a better understanding of the Treg cell-mediated effects in B-cell malignancies. We also anticipate that the clinical use of denileukin diftitox, an interleukin-2 and diphtheria toxin fusion protein, in combination with rituximab, an anti-CD20 monoclonal antibody, will inhibit Treg cells in B-cell lymphoma patients and will also deplete lymphoma B-cells in malignant lymph nodes thereby preventing further recruitment of Treg cells into areas of B-cell lymphoma. This treatment combination will lead to a novel therapeutic approach to modulating Treg cells that will result in clinical benefit for patients with B-cellNHL.